PROPOSAL SUMMARY The goal of my proposal is to understand the mechanisms regulating the self-renewal and lineage potential of hematopoietic stem cells (HSCs) and how HSC fate and function can be controlled by manipulation of epigenetic parameters. The context of my study is unique, because we have discovered a fetal, developmentally restricted HSC (drHSC) with unusual properties: the drHSCs are capable of long-term, multilineage reconstitution (LTMR) upon serial transplantation, but do not persist into adulthood during normal development. The ability of drHSCs to self-renew and persist is therefore induced upon transplantation. Further, while capable of generating all the ?traditional? hematopoietic cell types investigated to date, the drHSCs are lymphoid biased and have superior B1a cell reconstitution capacity compared to the co-existing fetal liver (FL) HSCs. Amazingly, the lymphoid bias and B1a capacity are retained over many months in serial transplantation experiments. The two core properties that define functional HSCs ? self-renewal and lineage potential ? are therefore uniquely regulated in drHSCs. I will leverage these unique properties to understand the molecular and epigenetic mechanisms that govern HSC fate and function, and how these mechanisms are both stable (as in the case of drHSC lineage potential) and dynamic (as in the case of induced persistence). I will also take advantage of my ability to isolate three distinct populations of HSCs: the drHSCs, co-existing fetal HSCs (fHSCs), and adult HSCs (aHSCs). Together, this will enable me to ask fundamental questions in HSC biology from a unique perspective and with novel strategies: How is long-term persistence of drHSCs induced upon transplantation, whereas their lineage bias is retained? Is lineage potential exclusively a loss-of-function phenomenon or do adult HSCs have differentiation capabilities that fetal HSCs lack? If so, how are these gained? How can one HSC population be ?reprogrammed? into another HSC subtype? I propose to pursue these questions by assessing epigenetic and transcriptome dynamics (Aim 1), and by functionally manipulating HSC potential using CRISPRi mouse models (Aim 2). Outcomes from this proposal will be provide fundamental proof-of-concept experiments for HSC manipulation with the long-term goal of applying these tools to increase the efficacy of stem cell transplant therapies.